The use of recombinant DNA technology has made possible the large scale production in fermentation facilities of proteins which would otherwise have to be isolated from natural sources. In the application of this technology, a microorganism such as a bacterium is transformed with a replicable expression vector containing a gene which codes for a protein not normally produced by the bacterial host, i.e., a heterologous protein. The gene for the heterologous protein is operably linked to a regulatory sequence of DNA, including a transcription promoter, which is capable of directing expression of the gene in the bacterial host. Due to the universality of the genetic code, the bacterial host is capable of transcribing the heterologous gene into messenger RNA (mRNA) and then translating the mRNA into protein having the amino acid sequence of the heterologous protein.
Over the last several years, a great deal of research in the recombinant DNA field has focused on methods for improving the efficiency with which transformed microorganisms can be made to express heterologous genes in order to improve yields of desired products. Efforts at improving levels of heterologous gene expression have been primarily directed at manipulation of "upstream" DNA components--that is, DNA sequences which precede the heterologous gene in the expression vector and affect the frequency and efficiency of transcription--and at controlling post-translational events which affect product stability or recovery. The latter approach is exemplified by efforts to develop host-vector combinations which effect secretion of the translated heterologous protein through the cell wall of the host cell and into the surrounding medium where it is less subject to degradative attack by proteases produced in the host cell.
While considerable improvements have been made in obtainable levels of gene expression and product recovery, the art continues to seek methods by which heterologous genes can be expressed and the expression products recovered in increased amounts.